PD-L1 upregulation in TAMs could be IFN-γ independent
(A–C) PBMCs from patients with BC were rested and stimulated with IFN-γ. Representative flow plots showing IFN-γ (50 ng/mL for 15 min) induced phosphorylation of STAT1 (pY701) in peripheral monocytes (A) from patients with BC. IFN-γ signaling response (n = 28) (B) and levels of IFN-γR1 (n = 20) (C) were compared between PD-L1–/lo vs. PD-L1+/hi monocytes.
(D and E) Rested PBMCs from patients with BC were stimulated with IFN-γ at 0.2, 1, 5, or 25 ng/mL for 16 h. Levels of PD-L1 and IFN-γR1 on monocytes or T cells are shown in the representative flow plots (D) and compared (n = 6) (E). One-way ANOVA.
(F–H) Single-cell suspensions from freshly prepared primary breast tumors were stimulated with IFN-γ. Representative flow plots (F) showing IFN-γ-induced pSTAT1 (G) and levels of IFN-γR1 (H) between PD-L1–/lo vs. PD-L1+/hi TAMs from untreated primary breast tumors (n = 8).
(I and J) Multiplex immunofluorescence staining (I) and quantification of IFN-γR1+ PD-L1+/− cells (J) from untreated primary breast tumor tissues (n = 8). Scale bars, 100 μm.
(K–M) Representative flow plots showing PD-L1 expression (K) and IFN-γ-induced pSTAT1 (L) and levels of IFN-γR1 (M) between PD-L1–/lo vs. PD-L1+/hi breast cancer cells (n = 8). Paired t test. ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. Shown are mean ± SEM.